Silencing SMYD3 in hepatoma demethylates RIZI promoter induces apoptosis and inhibits cell proliferation and migration

Figure 1 Enhanced expression of SMYD3 in HCC. A: The expression of SMYD3 genes was determined by RT-PCR and GAPDH served as an internal control; B: Representative images of immunohistochemical staining of accumulated SMYD3 protein in HCC tissue, but not in peri-cancerous tissue.

Figure 2 RNAi specifically inhibits SMYD3 expression in HCC cell lines. A: Schematic drawing of the Pgenesil-1 vector; B: The predicted secondary structures of the Pgenesil-1-s1 and Pgenesil-1-s2 transcripts target SMYD3 are shown; C: 48h after transfection, Western blot analysis showed the inhibitory effect of plasmids expressing SMYD3 shRNAs in HepG2 and SMMC-7721 cells.

Figure 3 Inhibition of SMYD3 reduces hepatoma cell proliferation (MTT assays).

Figure 4 Inhibition of SMYD3 reduces hepatoma cell migration. A: Representative cells (blue) invade through Matrigel and membrane; B: The percentage of cells successfully passing through the matrigel and membrane (P < 0.01).

Figure 5 Inhibition of SMYD3 promotes apoptosis in hepatoma cells upon serum deprivation. A: FCM analysis of HepG2 cells 48 h after transfection (P < 0.01). B: TUNEL assay of cell apoptosis with siRNA suppression of SMYD3 expression in HepG2 (brown staining cells) (P < 0.01).

Figure 6 Inhibition of SMYD3 de-methylates RIZ1 promoter CpG islands and reexpresses RIZ1 in hepatoma Cells. A: Methylation specific PCR (MSP) assay of RIZ1 promoter. M: Methylated; U: Unmethylated; B: RIZ1 mRNA expression, GAPDH served as internal control (RT-PCR).